Determinação de glifosato e AMPA em urina humana empregando o método dilute and shoot e UHPLC-MS/MS

• Ano de defesa: 2026
• Autor(a) principal: Torri, Júlia Maria Druzian
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Santa Maria; Brasil; UFSM; Programa de Pós-Graduação em Química; Centro de Ciências Naturais e Exatas
• Palavras-chave em Português: Glifosato; AMPA; Precipitação de proteínas; Urina; Glyphosate; Protein precipitation; Urine; Chromatography; CNPQ::CIENCIAS EXATAS E DA TERRA::QUIMICA
• Link de acesso: http://repositorio.ufsm.br/handle/1/38805

Pesticides are widely employed in agriculture for pest and weed control; however, their intensive use raises concerns due to the negative impacts they may cause on human health and the environment. Residues of these substances may persist in food, impair soil fertility, and reach groundwater, thereby compromising aquatic ecosystems. Chronic exposure to these compounds is associated with serious risks, including cancer, endocrine disorders, neurological damage, and developmental alterations in children. In this context, glyphosate stands out as the most widely used herbicide worldwide, due to its effectiveness in controlling invasive plants and its broad application in different agricultural crops. Among the biological matrices used to monitor such exposure, urine proves particularly useful for evidencing the excretion of toxic substances. In light of this, the present study evaluated the use of different analytical strategies such as protein precipitation, liquid–liquid extraction (LLE), salt-assisted liquid–liquid extraction (SALLE), solid-phase extraction (SPE), derivatization with FMOC-Cl, and dilute-and-shoot. These techniques were assessed for sample preparation aimed at the determination of glyphosate and its main metabolite, AMPA, in human urine using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). However, conventional sample preparation methods did not effectively remove matrix proteins. Thus, derivatization with FMOC-Cl was discarded, and the development was directed toward a direct HILIC-based method capable of overcoming the low selectivity observed. The validated analytical method, which showed the best performance, was applied to 250 μL of human urine transferred into a 2 mL polypropylene tube, to which 250 μL of 1 mmol L⁻¹ EDTA were added in order to complex metal cations and avoid interference in the determination. Subsequently, 500 μL of acetonitrile were added to aid in protein precipitation. The samples were then subjected to centrifugation (15,000 rpm/10 min/0 °C) and filtration (PTFE, 0.22 μm). Accuracy (recovery) results ranged from 74 to 108% at levels of 0.25, 0.5, 1.25, and 2.5 μg L⁻¹, all with relative standard deviations below 20%. Calibration curves prepared in the matrix extract showed linearity from 0.25 to 10 μg L⁻¹. The method exhibited a significant matrix effect for AMPA (-80%). Limits of detection (LOD) for glyphosate and AMPA were 0.30 μg L⁻¹, and the limits of quantification (LOQ) were 1 μg L⁻¹ for both analytes. After validation, the method was applied to 35 human urine samples collected from farmers in municipalities of Rio Grande do Sul State (Brazil), showing 1 sample ≤ LOQ for glyphosate and 8 samples ≤ LOQ for AMPA. The proposed method proved to be a suitable tool for monitoring glyphosate and AMPA in human urine samples.